Lactobacillus acidophilus R026 is unique among bacteria in its dependence upon four deoxynucleoside kinase activities for the synthesis of the deoxynucleoside triphosphate precursors of DNA. The products of these essential activities are kept in balance by combinations of end-product and substrate-level modulations. Three of the four activities appear to be organized into two overlapping pairs of associated activities, as evidenced by their kinetic interactions and their co-purification. The pairs observed are dCyd/dAdo and dGuo/dAdo phosphorylating species. The fundamental issues to be explored in this study are: 1) whether or not these paired activities are examples of multi-functional proteins, and 2) how a number of catalytic and regulatory sites are organized and interact on a relatively small protein molecule. The first question will be attached by use of a battery of affinity chromatography columns designed to resolve those activities not residing on a common protein molecule, followed by molecular weight determinantions of isotopically labeled subunits. In addition, mutants lacking one or more of these activities are to be selected from a wild-type strain. The second question will be probed using a combination of kinetics, active site-directed reagents, chemical modification of the protein and selective proteolysis, to determine which sites are overlapping or shared, whether modulation is achieved by effectors being bound to catalytic sites, or if distinct regulatory sites or subunits exist.